Retracted: In vivo and in vitro effects of microRNA-221 on hepatocellular carcinoma development and progression through the JAK–STAT3 signaling pathway by targeting SOCS3

Hepatocellular carcinoma (HCC), as the third leading cancer-caused deaths, prevails with high mortality, and affects more than half a million individuals per year worldwide. A former study revealed that microRNA-221 (miR-221) was involved in cell proliferation of liver cancer and HCC development. The current study aims to evaluate whether miR-221 targeting SOCS3 affects HCC through JAK–STAT3 signaling pathway. A series of miR-221 mimic, miR-221 inhibitor, siRNA against SOCS3, and SOCS3 plasmids were introduced to SMMC7721 cells with the highest miR-221 expression assessed. The expression of JAK–STAT3 signaling pathway–related genes and proteins was determined by Western blot analysis. Cell apoptosis, viability, migration, and invasion were evaluated by means of flow cytometry, 3-(4,5-dimethylthiazol-2-yl)-2,5 diphenyl tetrazolium bromide, and transwell assays, respectively. HCC xenograft in nude mice was performed to measure HCC tumor growth. miR-221 was found to be highly expressed but SOCS3 was poorly expressed in HCC tissues. miR-221 expression was correlated with lymph node metastasis (LNM) and tumor node metastasis (TNM) of HCC, and SOCS3 expression was correlated with LNM, differentiation and TNM of HCC. SOCS3 is a target gene of miR-221. MiR-221 mimic or si-SOCS3 exposure was found to induce cell viability, migration, and invasion, and reduce apoptosis. MiR-221 inhibitor was observed to have inhibitory effects on HCC cell proliferation, migration, and invasion. Moreover, the expression of JAK–STAT3 signaling pathway was suppressed by miR-221 inhibitor. Downregulated miR-221 expression could promote its target gene SOCS3 to inhibit the proliferation, invasion and migration of HCC cells by repressing JAK–STAT3 signaling pathway.     

Retracted: MicroRNA-520a-3p suppresses epithelial–mesenchymal transition,invasion, and migration of papillary thyroid carcinoma cells via the JAK1-mediated JAK/STAT signaling pathway

Papillary thyroid cancer (PTC) is a kind of thyroid cancer and frequently presents with epithelial–mesenchymal transition (EMT). MicroRNAs (miRNAs) were previously reported to be associated with PTC. Thus, this study aims to define the role of microRNA-520a-3p (miR-520a-3p) in PTC through the JAK/STAT signaling pathway by targeting JAK1. The PTC and normal thyroid tissues of 137 PTC patients were collected. First of all, the expression pattern of miR-520a-3p, JAK1, JAK2, STAT3, E-cadherin, and vimentin in PTC was identified. The relationship between miR-520a-3p and JAK1 was predicted and analyzed. And a series of miR-520a-3p mimic or inhibitor, or siRNA JAK1 introduced into PTC cells were applied to examine the effect of miR-520a-3p on PTC cell viability, migration, invasion, cell cycle, apoptosis, and EMT. Meanwhile, the regulatory effect of miR-520a-3p and JAK1 on the JAK/STAT signaling pathway was also determined. The expression of JAK1, JAK2, STAT3, and vimentin increased yet miR-520a-3p and E-cadherin decreased in PTC tissue. JAK1 was negatively regulated by miR-520a-3p. Functionally, EMT induction was prevented by miR-520a-3p upregulation through downregulating JAK1. When upregulating miR-520a-3p or silencing JAK1 in PTC cells, PTC cell viability, migration, and invasion were inhibited yet cell apoptosis promoted with cells arrested at G1 phase, indicating that miR-520a-3p prevented PTC progression by downregulating JAK1. Moreover, miR-520a-3p elevation or JAK1 inhibition inactivated the JAK/STAT signaling pathway. Collectively, miR-520a-3p prevents cancer progression through inactivating the JAK/STAT signaling pathway by downregulating JAK1 in PTC.     

Convallatoxin promotes apoptosis and inhibits proliferation and angiogenesis through crosstalk between JAK2/STAT3 (T705) and mTOR/STAT3 (S727) signaling pathways in colorectal cancer

BackgroundAberrant activation of STAT3 is frequently encountered and promotes survival, cellular proliferation, migration, invasion and angiogenesis in tumor cell. Convallatoxin, triterpenoid ingredient, exhibits anticancer pharmacological properties.PurposeIn this work, we investigated the anticancer potential of convallatoxin and explored whether convallatoxin mediates its effect through interference with the STAT3 activation in colorectal cancer cells.MethodsIn vitro, the underlying mechanisms of convallatoxin at inhibiting STAT3 activation were investigated by homology modeling and molecular docking, luciferase reporter assay, MTT assay, RT-PCR, Western blotting and immunofluorescence assays. Changes in cellular proliferation, apoptosis, migration, invasion and angiogenesis were analyzed by EdU labeling assay, colony formation assay, flow cytometry assay, wound-healing assay, matrigel transwell invasion assay and tube formation assays. And in vivo, antitumor activity of convallatoxin was assessed in a murine xenograft model of HCT116 cells.ResultsConvallatoxin decreased the viability of colorectal cancer lines. Moreover, convallatoxin reduced the P-STAT3 (T705) via the JAK1, JAK2, and Src pathways and inhibited serine-727 phosphorylation of STAT3 via the PI3K-AKT-mTOR-STAT3 pathways in colorectal cancer cells. Interestingly, we discovered the crosstalk between mTOR and JAK2 in mTOR/STAT3 and JAK/STAT3 pathways, which collaboratively regulated STAT3 activation and convallatoxin play a role in it. Convallatoxin also downregulated the expression of target genes involved cell survival (e.g., Survivin, Bcl-xl, Bcl-2), proliferation (e.g., Cyclin D1), metastasis (e.g., MMP-9), and angiogenesis (e.g., VEGF). Indeed, we found that convallatoxin inhibited tube formation, migration, and invasion of endothelial cells, and inhibited the proliferation. Finally, in vivo observations were confirmed by showing antitumor activity of convallatoxin in a murine xenograft model.ConclusionThe result of the current study show that convallatoxin promotes apoptosis and inhibits proliferation and angiogenesis through crosstalk between JAK2/STAT3 (T705) and mTOR/STAT3 (S727) signaling pathways in colorectal cancer cells and indicate that convallatoxin could be a valuable candidate for the development of colorectal cancer therapeutic.     

基于JAK2/STAT3信号通路探讨白藜芦醇对骨肉瘤MG-63细胞凋亡及迁移的影响

摘要 目的：研究白藜芦醇(RES)通过蛋白酪氨酸激酶2/信号转导子与激活子3（JAK2/STAT3）信号通路对人骨肉瘤体外细胞株MG-63细胞凋亡、侵袭和迁移的影响。方法：体外培养MG-63细胞,以不同浓度的RES作用于MG-63细胞。Annexin V-FITC/PI双染流式细胞术检测不同时间和不同浓度的RES对MG-63细胞凋亡的影响。划痕实验和Transwell实验检测不同时间和不同浓度的RES对MG-63细胞侵袭和迁移能力的影响。免疫印迹实验检测不同时间和不同浓度的RES对MG-63细胞磷酸化蛋白酪氨酸激酶2（p-JAK2）、磷酸化信号转导子与激活子3（p-STAT3）、凋亡相关蛋白B淋巴细胞瘤-2（Bcl-2）、Bcl-2家族促凋亡蛋白（Bax）及基质金属蛋白酶（MMP）-2、MMP-9表达的影响。结果：RES浓度越高,时间越久,MG-63细胞凋亡率越高（P<0.05）。RES浓度越高,MG-63细胞迁移和侵袭能力越弱(P<0.05)。RES处理MG-63细胞后其p-JAK2、p-STAT3、Bcl-2以及MMP-2、MMP-9的表达明显降低,而Bax蛋白表达明显升高,且p-JAK2、p-STAT3、Bax、Bcl-2以及MMP-2、MMP-9的表达水平变化具有RES浓度依赖性（P<0.05）。结论：RES可能通过调控JAK2/STAT3信号通路促使人骨肉瘤MG-63细胞凋亡,并抑制MG-63细胞侵袭和迁移。     

miR-155靶向SOCS1对骨肉瘤Saos2细胞的增殖、侵袭和迁移能力的影响

目的:探讨microRNA-155(miR-155)对骨肉瘤Saos2细胞增殖、侵袭和迁移的影响以及其作用机制。方法:利用实时荧光定量(qRT-PCR)实验检测miR-155在正常成骨细胞与骨肉瘤Saos2细胞中的表达水平,以及miR-155-mimic、miR-155-inhibitor的转染效率。采用CCK-8实验检测细胞的增殖能力,Transwell实验和划痕实验分别检测Saos2细胞的侵袭和迁移能力,Western blot检测细胞内的STAT3磷酸化水平以及SOCS1表达水平,双荧光素酶报告基因实验进行靶基因验证。结果:miR-155在骨肉瘤Saos2细胞中表达明显高于正常成骨细胞(P0.001)。在分别转染miR-155-mimic和miR-155-inhibitor后,Saos2细胞内miR-155表达水平明显上调和下降(P0.001)。过表达miR-155可促进Saos2细胞增殖、侵袭和迁移,降低SOCS1的蛋白水平,上调STAT3的磷酸化水平,差异均具有统计学意义。相反,降低miR-155水平可抑制Saos2细胞的增殖、侵袭和迁移能力,差异均具有统计学意义。结论:骨肉瘤Saos2细胞中高表达的miR-155可以通过抑制SOCS1表达来激活STAT3信号通路进而促进细胞的增殖、侵袭和迁移,因此,靶向抑制miR-155表达可以作为潜在治疗骨肉瘤的途径。     

CFTR activation suppresses glioblastoma cell proliferation,migration and invasion

The aim of this study was to investigate the function of Cystic fibrosis transmembrane conductance regulator (CFTR) in human glioblastoma (GBM) cells. Data dining results of the Human Protein Atlas showed that low CFTR expression was associated with poor prognosis for GBM patients. We found that CFTR protein expression was lower in U87 and U251 GBM cells than that in normal humane astrocyte cells. CFTR activation significantly reduced GBM cell proliferation. In addition, CFTR activation significantly abrogated migration and invasion of GBM cells. Besides, CFTR activator Forskolin treatment markedly reduced MMP-2 protein expression. These effects of CFTR activation were significantly inhibited by CFTR inhibitor CFTRinh-172 pretreatment. Our findings suggested that JAK2/STAT3 signaling was involved in the anti-glioblastoma effects of CFTR activation. Moreover, CFTR overexpression in combination with Forskolin induced a synergistic anti-proliferative response in U87?cells. Overall, our findings demonstrated that CFTR activation suppressed GBM cell proliferation, migration and invasion likely through the inhibition of JAK2/STAT3 signaling.     

沉默信息调节因子1通过诱导miR-221/222表达促进细胞自噬

微小RNA（microRNAs, miRNAs,）是一类强大的基因表达调控子,可在转录及转录后水平负调控靶基因的表达来参与生物学过程。沉默信息调节因子1 (silent information regulator1, SIRT1)底物众多,可通过去乙酰化作用参与多种细胞生命活动进程。尽管如此,SIRT1与非编码RNA如miRNA的表达调控关系仍有待深入研究。本文利用荧光定量PCR 检测发现,SIRT1与miR-221和miR-222的表达呈正相关：干扰SIRT1后,miR-221/222呈低水平表达;而过表达SIRT1则促进miR-221/222的表达。将miR-221/222基因簇启动子区序列插入pEZX-GA01构建双荧光素酶报告载体,与SIRT1过表达质粒或干扰序列共转至细胞。结果显示,SIRT1可显著提高miR-221/222启动子区活性,提示SIRT1可在转录水平调节miR-221/222的表达。进一步运用Western 印迹研究发现,在HEK293细胞中过表达miR-221/222可促进细胞的自噬能力,而抑制miR-221/222的表达可减弱自噬。此外,过表达SIRT1的同时抑制miR-221/222 的表达可减弱SIRT1的自噬诱导作用。综上所述,SIRT1可通过诱导miR-221/222的表达促进细胞自噬,其具体作用机制有待进一步探讨。     

MicroRNA-294 promotes cellular proliferation and motility through the PI3K/AKT and JAK/STAT pathways by upregulation of NRAS in bladder cancer

In our study we examined the role of microRNA-294 (miR-294) in bladder cancer and related mechanisms. Realtime polymerase chain reaction (RT-PCR) was performed to determine the expression level of miR-294. Western blot was used to determine the expression of NRAS, mainly factors in the PI3K/AKT and JAK/STAT pathways. Cell counting kit8 assay, clonogenic assay, wound-healing assay, transwell and flow cytometry were used to explore, respectively, cell proliferation, survival, migration, invasion, and apoptosis of bladder cancer cell line T24. The expressions of miR-294 in bladder cancer cells including J82, HT1376, T24, and SW780 were significantly increased compared to those in human bladder epithelium cells (both HCV29 and SV-HUC-1). The proliferation rate, surviving fraction, migration, and invasion of T24 cells in miR-294 mimetic transfected group were significantly increased, while they were significantly decreased by miR294 inhibitor transfection. Moreover, miR-294 suppression could increase the apoptotic rate of T24 cells. In addition, drug resistance of T24 cells to cisplatin was increased in miR-294 mimetic-treated group, while it was decreased by miR-294 inhibitor compared to empty control. Overexpression of miR-294 could upregulate NRAS expression in T24 cells and activate PI3K/AKT and JAK/STAT pathways. We found that miR-294 expression was positively related with proliferation and motility of T24 cells. Moreover, miR-294 suppression could promote the sensitivity of T24 cells to cisplatin. We also found miR-294 could upregulate NRAS and activate the PI3K/AKT and JAK/STAT pathways in T24 cells.     

Downregulation of microRNA-92b-3p suppresses proliferation,migration, and invasion of gastric cancer SGC-7901 cells by targeting Homeobox D10

To investigate the effect and mechanism of microRNA-92b-3p (miR-92b-3p) targeting Homeobox D10 (HOXD10) on proliferation, migration, and invasion of gastric cancer, we detected t he expression of miR-92b-3p and HOXD10 in SGC-7901 cells. The effects of miR-92b-3p or HOXD10 on proliferation, migration, invasion, and matrix metalloproteinase (MMP)-2/9 expression in SGC-7901 cells were measured by the Cell Counting Kit-8 assay, Transwell assay, and Western blot, respectively. The results showed that miR-92b-3p expression was increased, and HOXD10 expression was decreased in SGC-7901 cells, compared with human normal gastric epithelial cells GES-1. Functional experiments demonstrated that cell proliferation, migration, invasion, and expression of MMP-2/9 in SGC-7901 cells were significantly inhibited by miR-92b-3p silencing and HOXD10 overexpression. Moreover, HOXD10 was a potential target gene of miR-92b-3p as evidenced by the TargetScan software and double luciferase reporter assay. In the rescue experiment, knockdown of HOXD10, accompanied by higher expression of MMP-2/9, could significantly eliminate the inhibitory effects of miR-92b-3p silencing on cell proliferation, migration, and invasion. In conclusion, miR-92b-3p is highly expressed in gastric cancer SGC-7901 cells, and interfering with its expression might inhibit SGC-7901 cell proliferation, migration, and invasion via downregulating MMP-2/9 expression and targeting HOXD10.     

Berberine Inhibits Invasion and Metastasis of Colorectal Cancer Cells via COX-2/PGE2 Mediated JAK2/STAT3 Signaling Pathway

Berberin, extracted from Chinese herbal medicine Coptis chinensis, has been found to have anti-tumor activities. However, the underlying mechanisms have not been fully elucidated. Our current study demonstrated that berberin inhibited the in vitro and in vivo growth, migration/invasion of CRC cells, via attenuating the expression levels of COX-2/PGE₂, following by reducing the phosphorylation of JAK2 and STAT3, as well as the MMP-2/-9 expression. We further clarified that an increase of COX-2/PGE₂ expression offset the repressive activity of Berberin on JAK2/STAT3 signaling, and a JAK2 inhibitor AZD1480 blocked the effect of COX-2/PGE₂ on MMP-2/-9 expression. In summary, Berberin inhibited CRC invasion and metastasis via down-regulation of COX-2/PGE₂- JAK2/STAT3 signaling pathway.     

Curcumin blocks small cell lung cancer cells migration, invasion, angiogenesis, cell cycle and neoplasia through Janus kinase-STAT3 signalling pathway

Curcumin, the active component of turmeric, has been shown to protect against carcinogenesis and prevent tumor development. However, little is known about its anti-tumor mechanism in small cell lung cancer (SCLC). In this study, we found that curcumin can inhibit SCLC cell proliferation, cell cycle, migration, invasion and angiogenesis through suppression of the STAT3. SCLC cells were treated with curcumin (15 μmol/L) and the results showed that curcumin was effective in inhibiting STAT3 phosphorylation to downregulate of an array of STAT3 downstream targets ,which contributed to suppression of cell proliferation, loss of colony formation, depression of cell migration and invasion. Curcumin also suppressed the expression of proliferative proteins (Survivin, Bcl-X(L) and Cyclin B1), and invasive proteins (VEGF, MMP-2, MMP-7 and ICAM-1). Knockdown of STAT3 expression by siRNA was able to induce anti-invasive effects in vitro. In contrast, activation of STAT3 upstream of interleukin 6 (IL-6) leads to the increased cell proliferation ,cell survival, angiogenesis, invasion, migration and tumor growth. Our findings illustrate the biologic significance of IL-6/JAK/STAT3 signaling in SCLC progression and provide novel evidence that the pathway may be a new potential target for therapy of SCLC. It was concluded that curcumin is a potent agent in the inhibition of STAT3 with favorable pharmacological activity,and curcumin may have translational potential as an effective cancer therapeutic or preventive agent for SCLC.